Purpose :
Sorsby Fundus Dystrophy (SFD) is an autosomal dominant disorder that can result in central vision loss in early adulthood. Mutations in the TIMP3 gene are found in patients with SFD, but the pathogenesis remains unclear. In order to better understand the disease process, we determined the protein expression profile of retinal pigment epithelial (RPE) cells derived from an individual with SFD as compared to normal controls.

Methods :
Induced pluripotent stem cells (iPSCs) were generated from an individual with an S181C TIMP3 mutation and from unaffected control individuals. RPE cell lines were differentiated from at least three distinct iPSC clones from each individual and cultured to maturity (4 wks). RPE were analyzed using isobaric tag for absolute and relative quantification (iTRAQ) to develop a protein expression profile. The SFD-RPE and control-RPE samples were pooled separately, digested with porcine trypsin, and labeled with iTRAQ 4-plex reagents. All samples were then pooled together and fractionated with liquid chromatography followed by tandem mass spectrometry to produce a peptide sequence tag. This tag was searched against the proprietary peptide database Proteome Discoverer using a false discovery rate of 1% to identify the peptide and the corresponding parent protein. We then identified proteins that were expressed at either higher than two-fold or less than one-half-fold between the SFD and control samples. Western blot was used to confirm differences in protein expression.

Results :
We identified 6953 total proteins, 16 of which were present with increased abundance in SFD compared to control, and 9 of which were present with decreased abundance in SFD compared to control. Interestingly, several of the proteins with increased abundance in SFD were found to belong to the alpha and beta crystalline category and several were found to be involved in glutathione metabolism. The increased abundance of GSTA1 was confirmed with Western Blot.

Conclusions :
Several proteins with increased expression in SFD-derived RPE are involved in glutathione metabolism, which plays a role in cell protection against oxidative stress. A key protein involved in glutathione metabolism was confirmed using Western Blot. Our results suggest that glutathione metabolism may play a role in the pathogenesis of SFD. Further studies will be needed to determine the role of oxidative stress in Sorsby Fundus Dystrophy.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.